Cronobacter sakazakii, a globally prevalent dairy-borne pathogen with frequent pollution accidents worldwide, exhibited strong resistance to environmental stress, thus it was of great significance to develop novel antimicrobial agents against C.sakazakii. Thanatin (Tn), a promising antimicrobial peptide targeting the cell outer membrane, consists of 21 amino acids with a β-hairpin structure.Tn has been shown to exhibit bactericidal activity against Escherichia coli, yet there was a lack of reports regarding its antimicrobial effects against C.sakazakii.On the basis of this point, Tn was overexpressed, and its antimicrobial effect against C.sakazakii was revealed for the first time.The recombinant strains of Pichia pastoris were constructed to produce codon-optimized Tn variants via heterologous expression systems driven by PAOX and PGAP promoters respectively.The bactericidal activity against C.sakazakii BAA894 of purified Tn was assessed.To elucidate the underlying mechanisms, the sensitivities to Tn of the wild-type of C.sakazakii BAA894 and its lipopolysaccharide-deficient strain BAA894ΔwaaC were compared.Results demonstrated that both the constitutive GS115/pGAPZα A-than-1 and the inducible GS115/pPICZα A-than-1 produced the active Tn, with levels at 24.34 mg/L and 28.03 mg/L, respectively.After purified by His-tag affinity chromatography, Tn at 80 mg/L showed a bacteriostatic efficacy of 98.49% against C.sakazakii BAA894, demonstrating potent antibacterial activity.In contrast, the inhibitory effect on the BAA894ΔwaaC was decreased to 91.01%, displaying a 7.48% reduction compared to BAA894.This differential efficacy strongly suggested that the target of Tn was related to the core polysaccharide and O-antigen components of lipopolysaccharide.The present study lay a foundation for development and optimization of innovative antimicrobial agents against C.sakazakii.
YU Xia
,
YANG Wenyu
,
ZHOU Yun
,
WANG Shuiping
,
HU Xiaoqing
. Heterologous expression of recombinant thanatin and its antibacterial activity against Cronobacter sakazakii[J]. Food and Fermentation Industries, 2025
, 51(16)
: 20
-25
.
DOI: 10.13995/j.cnki.11-1802/ts.042619
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